7. Example 2 Pathogenic RAF1 S257LΒΆ

The next variant is a missense RAF1 variant in the 7th exon and is shown as number 2 in the variant cards of the sidebar. This variant sits on the opposite end of the spectrum and will be determined to be a straightforward pathogenic variant.

First look at the Population tab which shows that this variant is novel in both gnomAD Exomes Frequency and 1000 Genomes Frequency tracks which allows the inclusion of the first moderate criteria, PM2. Next, move on to the Gene Impact tab.

Another moderate criteria PM1 (in the Gene Region and Mutation Profile section) is confirmed here since this variant is located in a mutational hotspot with no known benign variants. Furthermore, with a high missense Z-score shown in the Missense as Mechanism of Disease section, the gene has a low rate of benign variants and is known to have other missense variants which provide a common mechanism for disease. This combines to capture the first supporting evidence criteria, PP2. Moving down further to the Computation Evidence section, the second captured supporting evidence is PP3 since SIFT and Polyphen predict the variant damaging and PhyloP and GERP++ predict it to be conserved. This completes the Gene Impact tab, so we move on to the Studies tab.

PM1 scoring criteria

Figure 7-1: Analyzing the PM1 scoring criteria.

The last two criteria to assess are in the Studies tab section. Due to the presence of multiple other high-quality (2+ star rating) pathogenic variants in ClinVar, the strong PS1 criteria and moderate PM5 criteria can be included. This of course is contingent upon the basis of the literature review, of which is easily accessible with VSClinical. In the Clinical and Functional Studies section, this variant can be searched in Google, Google Scholar or PubMed from their corresponding buttons. Also the detailed assessments and citations of labs submitting this variant to ClinVar can be reviewed. Two publications are easily found that reference not only this specific variant, but also other pathogenic variants in the same amino acid. These papers can be cited in the final interpretation. After completing these classifications, return to the Classification tab.

At this point all of the criteria have been accounted for, the interpretation built, and the final classification presented, which is pathogenic following rule iii of the ACMG rules having: 1 strong, AND 3 moderate 2 supporting criteria.

ACMG classification summary

Figure 7-2: The ACMG Classification summary panel.

In summary, this has been determined to be a straight-forward pathogenic variant, so we want to include this in the final report, and specifically in the primary findings. So, towards the bottom of the Classification tab for this variant, we will select the Primary Findings check box, verify that Pathogenic is selected in the Classification drop-down menu, and then click in the For Disorder: box. This brings up a list of associated phenotypes based on the OMIM annotation source. Select Noonan Syndrome 5, and we will leave the Inheritance / Variant Type: as the default.

interpretation section

Figure 7-3: Filling in the Interpretation section.

Next, the Interpretation: for this variant can be manually entered, but we want to start with the Auto Interpretation which is added to the Interpretation: text box by clicking the Add to Interpretation button in the Auto Interpretation dialog.

add auto interpretation

Figure 7-4: Adding Auto Interpretation.

This variant can not be finalized by selecting Finalize below the Interpretation: text box, and selecting Save & Close on the finalize variant dialog that appears.

finalize variant

Figure 7-5: Finalizing the variant.

save and close variant

Figure 7-6: Save and close this variant.

For the next example, we will take a deeper look at a stop gain variant.